Uas gcamp6m

Manufactured by Bloomington Drosophila Stock Center

Sourced in United States

UAS-GCaMP6m is a genetically encoded calcium indicator that can be used to monitor neuronal activity in Drosophila. It consists of a Gal4-responsive upstream activating sequence (UAS) that drives the expression of the GCaMP6m calcium sensor protein.

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14 protocols using uas gcamp6m

Visualizing mitochondria and DCVs in motor neurons

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w;+;D42-Gal4, UAS-mitoGFP (Dr. Pallanck, University of Washington) and w;sp/Cyo;D42-Gal4, UAS-ANF-GFP (Dr. Saxton, UC Santa Cruz) were used to visualize GFP signals in mitochondria and dense core vesicles (DCVs), respectively, in motor neurons. UAS-SOD1 and UAS-SOD2 from the Bloomington Stock Center were crossed with w;+;D42-Gal4, UAS-mitoGFP to generate UAS-SOD1; D42-Gal4, UAS-mitoGFP and UAS-SOD1; D42-Gal4, UAS-mitoGFP. We used these two strains to perform the rescue experiments in Figs 1, 2 and 4. To examine calcium (Ca²⁺) levels in motor neurons, we received fly strains UAS-GCaMP6m from the Bloomington Stock Center, and generated UAS-GCaMP6m/+; D42-Gal4/+. To activate or down-regulate the JNK pathway, we used UAS-Hep^B2 (Bloomington Stock Center) and UAS-Bsk RNAi (VDRC), respectively. These fly strains were also crossed with the control strain, w;+;D42-Gal4, UAS-mitoGFP, to generate UAS-Hep^B2; D42-Gal4, UAS-mitoGFP and UAS-Bsk RNAi; D42-Gal4, UAS-mitoGFP. All flies were maintained in normal fly food at 25°C with 40%-60% humidity and a 12 hr light/dark cycle.

Liao P.C., Tandarich L.C, & Hollenbeck P.J. (2017). ROS regulation of axonal mitochondrial transport is mediated by Ca2+ and JNK in Drosophila. PLoS ONE, 12(5), e0178105.

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Comprehensive Fly Strain Database for Research

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We used the following fly strains: nSyb-Gal4 (gift from Shanhaz Lone), UAS-GCaMP6m (Bloomington #42748), UAS-H2B:RFP (gift from Boris Egger), GMR57BO4-Gal4 (Bl. #46355), Gr94a-gal4 (Bl. #57686), Gr21a-Gal4 (Bl. #23890), Gr22e-Gal4 (Bl. #57608), Gr66a-Gal4 (Bl. #57670), Gr33a-Gal4 (Bl. #57623), Gr59d-Gal4 (Bl.57653), Gr32a-Gal4 (Bl. #57622), Gr97a-Gal4 (Bl. #57687), Gr59e-Gal4 (Bl. #57655), UAS-myr:GFP (Bl. #32198), and UAS-TNTE (Bl. #28837). Gr43a lines: Gr43a-KI-Gal4 mutant, Gr43a-KI-GAL4; UAS-Gr43a and Gr43a-KI-GAL4; UAS-Cha7.4kb were kindly offered by Hubert Amrein.

Maier G.L., Komarov N., Meyenhofer F., Kwon J.Y, & Sprecher S.G. (2021). Taste sensing and sugar detection mechanisms in Drosophila larval primary taste center. eLife, 10, e67844.

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Drosophila Rearing and Genetic Lines

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Flies were raised on standard cornmeal food media at 25°C and 70% relative humidity under a 12:12-hour light: dark cycle. The “Cantonized” w¹¹¹⁸w (CS10) strain was used as a wild-type control. The MB-GAL80, UAS-GCaMP6m, and OK107-GAL4 flies were obtained from Bloomington Drosophila stock center. The UAS-eNpHR-YFP; UAS-eNpHR-YFP fly line was obtained from Ann-Shyn Chiang. The RNAi lines were obtained from the Vienna Drosophila RNAi Center or TRiP RNAi fly stocks. All RNAi lines from the Vienna Drosophila RNAi Center have been described previously[12 (link)]. The VT30604-GAL4, VT44966-GAL4, C739-GAL4, VT49246-GAL4, VT0765-GAL4, UAS-shi^ts, and tubP-GAL80^ts flies have been described[20 (link)].

Shyu W.H., Lee W.P., Chiang M.H., Chang C.C., Fu T.F., Chiang H.C., Wu T, & Wu C.L. (2019). Electrical synapses between mushroom body neurons are critical for consolidated memory retrieval in Drosophila. PLoS Genetics, 15(5), e1008153.

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Rearing Drosophila Flies in Standard Media

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Flies were reared at 22°C–25°C in standard cornmeal-dextrose media. Wild type flies used in the experiments were Canton-S (BL 64349) obtained from Bloomington stock center. Orco-GAL4 (BL 23292) and UAS-GCaMP6m (BL 42748) flies were also obtained from Bloomington stock center. Following is the composition of the media per 100 food vials: dextrose-100 gms, inactive dry yeast-50 gms cornmeal-70 gms, Drosophila agar-6 gms, propionic acid-6 mL, tegosept-12 mL, water-1025 mL.

Clark J.T., Ganguly A., Ejercito J., Luy M., Dahanukar A, & Ray A. (2022). Chemosensory detection of aversive concentrations of ammonia and basic volatile amines in insects. iScience, 26(1), 105777.

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Drosophila Husbandry and Genetic Manipulation

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Flies were kept in vials containing a standard medium made of yeast, corn, and agar at 25°C, 60% relative humidity, and on a 12 hr light-12-hr dark cycle. Virgin female flies were collected shortly after eclosion and kept in groups (25 flies per vial) on standard fly medium (ND, with 10% sucrose) or HSD (ND plus an additional 10% sucrose) for 4–6 days before experiments.
Fly strains used in the manuscript: nosNGT-GAL4: (#31777), Maternal-tubulin-Gal4 (#2318), Gr5a-GAL4 (#57592), and UAS-GCaMP6m (#42748) were obtained from the Bloomington Drosophila Stock Center at Indiana University; UAS-E(z) RNAi (#2831), UAS-Pcl RNAi (#1185), UAS-Su(var)3–9 RNAi (#3558), UAS-Rpd3 RNAi (#0695), and UAS-cad RNAi (#03,877.N) were from the Tsinghua Fly Center.

Yang J., Tang R., Chen S., Chen Y., Yuan K., Huang R, & Wang L. (2023). Exposure to high-sugar diet induces transgenerational changes in sweet sensitivity and feeding behavior via H3K27me3 reprogramming. eLife, 12, e85365.

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Drosophila Strains for Neuroscience Research

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Drosophila melanogaster strains were raised on standard cornmeal/agar medium supplemented with dry yeast at 25 °C with a 12 hr light/dark cycle. The wild type stock was a w¹¹¹⁸strain. The following stocks were obtained from Bloomington Stock Center: w¹¹¹⁸;dTrpA1¹ (BL26504), dTrpA1 RNAi1 (BL31384), w*;dTrpA1¹, dTrpA1-Gal4 (BL36922), w*; dTrpA1-Gal4 (BL 27593), UAS-RFP (BL 27391), UAS-GCaMP5 (BL 42037), UAS-GCaMP6m (BL 42748). The Gr66a-IRES-GFP vector was kindly provided by Kristin Scott, the NSyB-Gal4;UAS-GCaMP3 was obtained by Patrick Verstreken. The lines Gr66aGal4, UAS-dTrpA1(A);dTrpA1^ins,dTrpA1GAL4, UAS-dTrpA1(B);dTrpA1^ins,dTrpA1Gal4 and the dTrpA1 RNAi 2 were kindly provided by the laboratory of Paul Garrity.

Soldano A., Alpizar Y.A., Boonen B., Franco L., López-Requena A., Liu G., Mora N., Yaksi E., Voets T., Vennekens R., Hassan B.A, & Talavera K. (2016). Gustatory-mediated avoidance of bacterial lipopolysaccharides via TRPA1 activation in Drosophila. eLife, 5, e13133.

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Drosophila Husbandry and Genetic Manipulation

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Fly Genetics for Neural Calcium Imaging

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In this study, two stable fly strains carrying the Gal4 driver and UAS-GCaMP on the same chromosome were made by recombination: c164Gal4-UAS-GCaMP1.3 (on the 2^nd chromosome, abbreviated hereafter c164-GCaMP1.3, see Torroja et al., 1999 for c164-Gal4, a motor neuron driver) stock was used to carry out most of experiments, and nSynaptobrevinGal4-UAS-GCaMP6m (nSyb-GCaMP6m, nSyb-Gal4 is a pan-neuronal driver on the 3^rd chromosome) was used to confirm then results. The UAS-GCaMP1.3 stock was a generous gift from Drs. Yalin Wang and Yi Zhong of Cold Spring Harbor Laboratory, Cold Spring Harbor, NY (Nakai et al., 2001 (link); Wang et al., 2004 (link); Ueda and Wu, 2006 (link)). UAS-GCaMP6m was from the Bloomington stock center (Bloomington, IN, see also Chen et al., 2013 (link)). nSyb-Gal4 was a gift from Dr. Toshihiro Kitamoto of the University of Iowa. In addition, K⁺ channel mutational line eag¹Sh¹²⁰/Y; c164-GCaMP1.3 and Na⁺ channel mutational line para^bss1/Y; c164-GCaMP1.3 were created for analysis (cf. Xing and Wu, 2018 ). For simplification, eag¹Sh¹²⁰ is abbreviated as eag Sh, and para^bss1 is abbreviated as bss. OK371-mCD8GFP was constructed from OK371-Gal4 (2^nd) and UAS-mCD8GFP (2^nd) for the use in electrophysiological recording.

Xing X, & Wu C.F. (2018). Inter-relationships among physical dimensions, distal-proximal rank orders, and basal GCaMP fluorescence levels in Ca2+ imaging of functionally distinct synaptic boutons at Drosophila neuromuscular junctions. Journal of neurogenetics, 32(3), 195-208.

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Genetic Tools for Olfactory Research

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All flies were raised on standard cornmeal agar medium, under 60% humidity and a 12-h light/12-h dark cycle at 25 °C. The Or10a-Gal4 (BL9944), Or42b-Gal4 (BL9971), Or43a-Gal4 (BL9974), Or49b-Gal4 (BL9986), Or56a-Gal4 (BL9988 and 23896), Or82a-Gal4 (BL23125), Or85a-Gal4 (BL23133), Or92a-Gal4 (BL23140), UAS-Orco (BL23145), Orco^{1 (link)} (BL23129), Orco^{2 (link)} (BL23130), UAS-TNT (BL28837 and 28997), UAS-TNT (inactive, BL28844), UAS-GCaMP6m (BL42748 and 42750), UAS-GCaMP6f (BL42747), and UAS-H134R-ChR2 (BL28995) flies were from the Bloomington Stock Center. UAS-mCD8-GFP was a gift from Dr. Chris Potter at the Johns Hopkins University School of Medicine, USA. UAS-P2X₂ was a gift from Dr. Zuoren Wang at the Institute of Neuroscience, China. The flies have been backcrossed for seven generations to a laboratory w¹¹¹⁸ strain.

Cao L.H., Yang D., Wu W., Zeng X., Jing B.Y., Li M.T., Qin S., Tang C., Tu Y, & Luo D.G. (2017). Odor-evoked inhibition of olfactory sensory neurons drives olfactory perception in Drosophila. Nature Communications, 8, 1357.

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Drosophila Strain Collection and Acclimation

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The fly strains r4-GAL4 (#33832), elav-GAL4 (#8760), TRPA1-GAL4 (#27593), TRP-GAL4 (#36359), TRPL-GAL4 (#52274), R11F02-GAL4 (#49828), iav-GAL4 (#52273), TRPA1-AB-GAL4 (#67131), TRPA1-CD-GAL4 (#67133), UAS-TeTxLC (#28838) and UAS-GCaMP6m (#42748) were obtained from the Bloomington Drosophila Stock Center (Indiana, USA). Myo31DF-GAL4 (#112001) and tub-GAL4 (#108074) strains were obtained from Kyoto stock center (DGRC) (Kyoto Institute of Technology, Japan). w¹¹¹⁸ was provided from NIG-FLY stock center (National Institute of Genetics, Japan).
Fly stocks were raised on a corn/yeast/glucose medium containing 80 g brewer's yeast powder, 100 g glucose, 40 g cornmeal, and 7 g agar, 8.8 mL propionic acid, and 0.88 g butyl parahydroxybenzoate per 1 L water. Fly stocks were cultured at 25 °C with a 12-h light/12-h dark cycle. The strain w¹¹¹⁸ was used as a control. In cold acclimation assays, vials were placed in an incubator at 18 °C for 1 day (20–23 h).

Suito T., Nagao K., Takeuchi K., Juni N., Hara Y, & Umeda M. (2020). Functional expression of Δ12 fatty acid desaturase modulates thermoregulatory behaviour in Drosophila. Scientific Reports, 10, 11798.

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